Identifying the interactions between thymocytes and thymic stromal cells involved in shaping the T cell antigen receptor repertoire and defining the stromal cell populations mediating these processes are the long range goals of this research program. The first aim of this proposal will utilize immunoelectron microscopy to characterize the thymic distribution of selected TCR VBeta gene products within the context of several model systems established to examine intrathymic selection of cells expressing these molecules; alloantigen-mediated selection in normal mice, neonatal tolerance to environmental antigens, and selection in transgenic mice expressing transgene T cell antigen receptor molecules. The second aim will ultrastructurally identify the cell types that express Class II molecules in thymus tissue from Class II-MHC transgenic mice with different intrathymic patterns of transgene expression. These studies will provide a unique opportunity to correlate the expression of Class II products by defined populations of thymic stromal cells with the ability of those cells to affect thymic selection and may help resolve some of the apparently contradictory results obtained with these lines of mice. The third aim will address the hypothesis that Class II molecules or complexes of self- peptides and Class II molecules expressed by thymic epithelial cells possess epitopes unique from those expressed by hematogenous cells. These studies will examine the ability of monoclonal antibodies and alloreactive T cell hybridomas raised against thymic epithelial Class II molecules to recognize Class II molecules expressed by hematogenous cells. The final aim will test the hypothesis that the decline in T cell function and the increased incidence of autoimmune disease seen with advancing age is correlated with an age-related decline in the ability of the thymus to influence the TCR repertoire. The ability of thymuses from neonatal, young and old mice to positively and/or negatively select thymocytes expressing selected VBeta gene products will be analyzed by flow cytometric, molecular biological, immunohistochemical and morphometric approaches.